Project Summary The gut microbiota is a dense polymicrobial consortium housed in the mammalian gastrointestinal tract that participates in human development, health and disease. The taxonomic composition of the microbiota changes during the progression of many different intestinal diseases including inflammatory bowel disease and colorectal cancers. While the causes underlying disease-associated microbiota changes are unclear, diseased tissues exhibit altered production of mucosal polysaccharides, which serve as nutrients for the microbiota. We hypothesize that changes in mucosal polysaccharide identity and abundance facilitate alterations in the microbiota composition by favoring or disfavoring the expansion of various gut taxa that specialize in the utilization of various mucosal polysaccharides. However, identifying such changes in mucosal polysaccharide content and abundance is limited by the absence of rapid and efficient methods to monitor fractionated polysaccharide preparations extracted from human mucosal samples. Fortuitously, some microbiota members residing in the gut have evolved specialized sensor proteins that detect specific complex polysaccharide structures and direct rapid and dramatic changes in gene expression. These sensors can distinguish between compositionally similar yet structurally distinct polysaccharides and are activated even when their cognate ligand is present in relatively low abundance in a heterogenous polysaccharide mixture. Here, we harness the specificity of these polysaccharide sensors to develop arrayed reporter libraries that employ gut microbes to evaluate changes in mucosal polysaccharide content in heterogenous mixtures. This work will develop a high- throughput surveillance system to define the polysaccharide content of the gut mucosa, efficiently monitor fractionation of complex polysaccharide mixtures and assign bacterial gene content to the utilization of individual polysaccharides. Ultimately, this toolset will unveil how intestinal diseases promote changes in the microbiota composition and output, identify putative disease biomarkers and offer valuable insight into the inter-kingdom interactions governing the human gut microbiota.